Diagnosing Bernard-Soulier syndrome (BSS), a congenital hemorrhagic disorder of blood platelets, is complicated by the difficulty of separating the giant platelets from other blood cells to allow studies of platelet function and structure. We report on the use of three whole blood assays for diagnosing BSS. Whole blood platelet aggregation responses studied with an electrical impedance aggregometer were equivalent to those more laboriously obtained by using platelet-rich plasma prepared by unit gravity sedimentation and studied with an optical light transmittance aggregometer. Platelet aggregation responses were normal with adenosine diphosphate or collagen stimulation but absent with ristocetin or bovine plasma stimulation. Whole blood radioimmunoassay of platelet glycoprotein (GP) expression was performed by using iodinated murine monoclonal antibodies HP1-1D (anti-GP IIb/IIIa) and 6D1 (anti-GP Ib). After incubation with citrated whole blood, centrifugation was used to separate cell-bound antibody that was quantitated with a gamma counter. The patient's whole blood had a normal level of cell-bound GP IIb/IIIa but a substantially reduced level of cell-bound GP Ib (5% of normal mean). Whole blood smear immunocytochemical staining with monoclonal antibodies and qualitative analysis by light microscopy revealed a considerable reduction of GP Ib expression by the patient's giant platelets, whereas GP IIb/IIIa expression was normal. These data helped establish the diagnosis of BSS. We conclude that these three relatively simple assays of platelets in whole blood should be of particular value in the clinical laboratory differential diagnosis of patients with congenital thrombocytopenias and giant platelet syndromes.